Summary: Replication-competent neuroattenuated herpes simplex virus (HSV-1) vectors currently used in Phase 1 clinical trials behave differently in different cell types and under different physiological conditions. Thus, the risk associated with the use of a specific vector will differ from one clinical application to the next. To adequately evaluate what clinical data from one trial is relevant to another, further characterization is needed of 1) the cell types in which these neuroattenuated vectors replicate and 2) the cellular and viral factors important for the replication of these vectors in this restricted range of cell types. During my first year at CBER my laboratory has begun to address these long-term questions by assembling the equipment and reagents required for making viral mutants and examining the expression of viral proteins in different cell types in vitro. Another aspect of this project is the possibility that HSV-1 uses an alternate mechanism of protein translation--one controlled by an Internal Ribosome Entry Site (IRES)-- to circumvent cell- and virus-mediated protein synthesis shut down in virally infected cells. Only recently has a member of the herpes virus family (Kaposi's Sarcoma-associated Herpes Virus, KSHV) been reported to encode a protein regulated by an IRES. Like the neuroattenuated vectors in use in clinical trials, IRES-regulated protein expression can be dependent on the cell type and its physiological status. The ability of IRESs to operate in HSV-infected cells will be examined.